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Gene expression profiles at single cell level of nucleotides optimized WT or S449A PTPN22 in J.Lck WT or J.Lck Y192F Jurkat cells

ABSTRACT: Gene expression profiles at single cell level of nucleotides optimized WT or S449A PTPN22 in J.Lck WT or J.Lck Y192F Jurkat cells

PROVIDER: PRJNA1105053 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR28817182.fastq.gz	Fastqsanger.gz
	SRR28817183.fastq.gz	Fastqsanger.gz
	SRR28817184.fastq.gz	Fastqsanger.gz
	SRR28817185.fastq.gz	Fastqsanger.gz
	SRR28817186.fastq.gz	Fastqsanger.gz

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Project description:Protein tyrosine phosphatase nonreceptor type 22 (PTPN22) gene segregates with most autoimmune diseases; its risk allele encodes overactive PTPN22 phosphatases that alter B cell receptor (BCR) signaling potentially involved in the regulation of central B cell tolerance. To assess whether PTPN22 risk allele affects the removal of developing autoreactive B cells, we tested by ELISA the reactivity of recombinant antibodies isolated from single B cells from asymptomatic healthy individuals carrying one or two PTPN22 risk allele(s). We found that new emigrant/transitional and mature naive B cells from PTPN22 risk allele carriers contained high frequencies of autoreactive clones compared to non-carrier control donors. Hence, a single PTPN22 risk allele has a dominant effect on altering autoreactive B cell counterselection, suggesting that early B cell tolerance checkpoint defects precede the onset of autoimmunity. In addition, gene array experiments comparing mature naÃ¯ve B cells from healthy individuals carrying or not PTPN22 risk allele(s) revealed that the strength of association of PTPN22 for autoimmunity, second in importance only to the MHC, may not only be due to BCR signaling alteration but also to the regulation of other genes, which themselves have also been identified as involved in the development of autoimmune diseases. The PTPN22 risk allele is a single nucleotide change (cytidine to thymidine) at residue 1858, which results in a single amino acid substitution from arginine to tryptophan at position 620 of the PTPN22/Lyp protein. Data from mature naÃ¯ve B cell populations from patients carrying 1 or 2 PTPN22 T alleles and non-carrier patients were compared in order to characterize the impact of PTPN22 polymorphism on B cell physiology. RNA was extracted from batch-sorted CD19+CD10-CD21+CD27- conventional mature naive B cells using the Absolutely RNA microprep kit (Stratagene). 100-200 ng of RNA was obtained per sample, and the quality of the purified RNA was assessed by the Bioanalyzer from Agilent. Using the Ovation biotin system kit from Nugen, 30-50ng of RNA was amplified and labeled to produce cDNA. Labeled cDNA was hybridized on chips containing the whole human genome (Human Genome U133 Plus 2.0 from Affymetrix).

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Gene expression profiles at single cell level of nucleotides optimized WT or S449A PTPN22 in J.Lck WT or J.Lck Y192F Jurkat cells

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